The vulnerability of some women to develop major depressive disorder (MDD), which occurs more frequently in women than in men, is associated with hormonal fluctuations (monthly, pregnancy, postpartum, and menopause). We have shown recently, using the technique of in vivo chronoamperometry, that acute administration of ovarian hormones (estradiol benzoate (EB) or progesterone (P)) impacts the ability of selective serotonin reuptake inhibitors (SSRIs) to alter the function of what is widely considered their initial cellular target in brain - the serotonin transporter (SERT). In addition, EB but not P, when given acutely, blocked SERT function as demonstrated by a diminished clearance of exogenously applied serotonin. Thus, EB has two actions. On its own, it slows the clearance of 5-HT, an effect similar to that caused by SSRIs. One might speculate that estrogen is antidepressant based on such an effect. However, we also find estrogen to interfere with the ability of SSRIs to slow 5-HT clearance. Such an effect might be expected to compromise the antidepressant effects of SSRIs. Estrogen, then, seems to have two quite distinct effects. By contrast, progesterone only seems to inhibit the effect of SSRIs on 5-HT clearance. There appears to be some difference in the mechanism(s) mediating these two effects of estradiol. Evidence was obtained that these effects of estradiol are mediated via activation of membrane as well as nuclear estrogen receptors (ER), indicating a genomic as well as a non-genomic component to these effects. By contrast, the effect of progesterone seems to be mediated primarily by nuclear receptors. A principal goal of this proposal is to extend these studies using other measures, either neurochemical (in vivo microdialysis) or behavioral (forced swimming test (FST)), to see if additional types of evidence can be obtained showing that treatment with female sex hormones either influences SERT function and/or interferes with the ability of SSRIs to inhibit the SERT. In addition, to examine possible mechanisms underlying hormonal effects, experiments are planned to study the plasma membrane distribution of the SERT using biotinylation studies; to examine the role of specific ER subtypes; and to study possible involvement of brain-derived neurotrophic factor (BDNF), as we found its administration to mimic the effect of EB and P on the ability of SSRIs to inhibit the SERT. Although the primary focus is the hormone/SSRI interaction, some studies focus on the inhibitory effect of estrogen alone. Finally, in a rat model of hormone-induced pseudopregnancy and subsequent hormone withdrawal, we will study the effects of high hormone levels administered chronically and their withdrawal in the FST and on the phosphorylated state of TrkB as well as their influence on SSRI/SERT interactions. PUBLIC HEALTH RELEVANCE: By 2020, the World Health Organization has predicted that MDD will be the second leading cause of disease burden worldwide, with women in their reproductive years representing over two-thirds of that estimate since women are twice as likely to suffer from MDD as men. Our studies involve the interaction between ovarian steroids, the selective serotonin reuptake inhibitor (SSRI) class of antidepressants, and the serotonin transporter (SERT), in particular examining the mechanisms by which such steroids interfere with the ability of SSRIs to block the SERT. The results obtained could have important implications for understanding treatment non-response in some women.